Five days a
week, Charles "Chip" Hawkins plays God. He peers down into
a microscope and with his left hand twists a lever that
holds a mouse embryo in place. With his right hand, he
squeezes another lever, this one controlling a
one-micron-wide pipette that pokes the embryo, the product
of a coupling 12 hours ago. The sphere bunches up before
yielding to the pipette's prick, which injects a few
hundred strands of DNA into one half of the budding
nucleus. The embryo relaxes, as if instantly coming to
terms with this transformation of its nature, and returns
to being perfectly round.

Hawkins pushes it to the side, uses a suction device to
grab another, and repeats the process — up to 10 or
so injections a minute, 600 a day. Hawkins, manager of the
Johns Hopkins School of Medicine's Transgenic Core
Laboratory, and his staff of two spend weekdays nestled in
a stone-silent room high up and deep inside a maze-like
building on the Johns Hopkins medical campus, fulfilling
125 or so requests each year from Hopkins researchers to
create mice that will become the living tests of their
hypotheses.

Hawkins and crew will make transgenic mice — mice
with one gene specifically added to make them useful in
narrow areas of research — like the ones Hawkins
injects now for the lab of Michael K. Lee, a
neuropathologist at the School of Medicine. Lee will use
the mice to study Parkinson's disease. Hawkins' group might
also make a mouse from stem cells taken from a mouse
embryo, then grown in a lab and injected with DNA that will
switch off a specific gene. The modified stem cells are
then transferred to a new embryo to be carried by a
surrogate mother, who will give birth to mice that can be
bred with "normal" ones to create a so-called knockout
mouse (because the gene was "knocked out") so researchers
can deduce that gene's function.

"In 25 years,
80 percent of the
discoveries
we'll see will be traced directly back to the
research mouse,"
says Pardoll.

Once he's done, Hawkins will use a siphon to suck healthy
embryos — the ones not turning pale, which means
they're dying — into a pipette, and then move them
into a dish no bigger than a quarter. Later in the day, the
transgenic staff will implant them in female mice that have
made whoopie with vasectomized males, so their bodies will
release hormones. "The females have to think there is a
reason for them to be pregnant for this to work," Hawkins
says. In 19 or so days, the new mice will be born,
eventually growing to around 20 grams. The cost: about
$3,500 per mouse.

But one fully grown mouse is a mere blip on the continuum
of what has become a mouse-making industry. "With mice, you
spend several years breeding and figuring out what your
transgenic mouse is about and what it can do," says
Hawkins. Genetically enhanced mice can always be bred with
others to make different — and better —
research subjects. "Researchers will spend more time to
make sure they haven't inadvertently affected a gene they
didn't want to affect. Making the right mouse is an ongoing
process."

Welcome to the Cult of the Mouse.

Between 20 million and 30 million mice are used nationally
each year in biological and medical research, dwarfing the
numbers of other animals, including 40,000 monkeys, used to
further the aims of science annually. The federal
government, as well as university and other labs, "make"
many of those millions of mice specifically for a purpose.
The world's leading mouse factory, the Jackson Laboratory
in Bar Harbor, Maine, breeds 3,500 specialized strains of
mice, freezes their embryos and sperm, devises new ways to
implant genes, and provides mice to virtually every leading
research institution in the United States, including
Hopkins. Its 2007 revenue: $150 million. Last year, Jackson
shipped 2.5 million mice around the world. All of its
rodents descend from a handful collected 100 years ago by
the company's founder, a scientist and university president
named Clarence Cook "C. C." Little — either the
George Washington or Pied Piper of the modern scientific
research mouse, depending on how you look at it (see "A
Tiny History of a Wee Research Tool," page 36).

Humans, who
have long celebrated clever ways to kill the
skittish-but-intrusive rodent, have now cast their lot with
it. Rather than build a better mousetrap, we're trying to
build a better mouse. Ever since scientists learned how to
manipulate a mouse's genes in the 1980s — making it
even more suitable for ever-more specific research —
the house mouse, or Mus musculus, has become a
scientific powerhouse. The sequencing of the mouse genome
six years ago made the mouse a seemingly endless tool for
scientific exploration.

"In 25 years, 80 percent of the discoveries we'll see will
be traced directly back to the research mouse," says Drew
Pardoll, a professor of
oncology at the School of Medicine and a leading
researcher on the interrelationship between the immune
system and cancer. "The past 25 years have just been
incredible. The pace and breadth of research has just
exploded — I mean, like times 100. The mouse has
always been big. It's just gotten progressively bigger."

Inbred mice have been key. The standard issue of the mouse
industry — the eight basic strains of inbred mice
— are cheaply produced (unlike their genetically
rejiggered cousins) and have the advantage of
predictability: Like twins, they carry the same exact
genes. "The beauty of inbred mice is that because each one
is a carbon copy of another, you can make better
comparative studies," says Pardoll. "You can give five mice
one type of therapy, and five others another type, and see
which is more effective."

Pardoll uses his colony of 3,000 mice to develop and test
drug therapies — as has been done around the world
for decades — because they have several things in
common with humans. They're mammals like us. They have
immune systems that are 98 percent identical to ours and
genetic make-ups that are about 95 percent similar. In
other words, they translate well to human aims, such as the
drive to cure cancer.

"The mouse has provided us with an excellent way to model
cancer in a mammalian system," says Pardoll. "Every single
insight into how the cancer cell and the immune system
interact comes from research on the mouse."

Keyata Thompson with her murine charges in the Ross
Research Building.

In the past, such insights were achieved through
implanting human tumor cells in "nude mice" —
hairless rodents with unhealthy or knocked-out immune
systems. Tumors grew freely in nude mice, and researchers
could easily monitor the growth. But in the past seven
years or so, Pardoll says, mice have been genetically
engineered to grow their own tumors, which makes them
better subjects for research work and clinical studies that
can more precisely approximate the development of human
malignancies. For example, mice bred to develop prostate
tumors have allowed scientists to study a specific immune
system response to cancer cells created from their own
cells, from within the same biological system. Scientists
have been able to devise ways to combine antibodies and
vaccines to fight breast and pancreatic cancer and multiple
myeloma in mice, and are now translating that work into
human clinical trials.

Hopkins researchers from other medical specialties have
made gains by exploiting the species' genome to genetically
engineer mice to help scientists determine the series of
genes that cause Down syndrome. They've made several types
of mice that mimic human schizophrenia. They've tested
drugs and compounds created to prevent skin cancer, cure
malaria, and reverse a potentially fatal weakening of
arteries in patients with Marfan syndrome. Several
neurologists and brain scientists oversee a one-of-a-kind
lab at the Homewood campus that develops and tests mice for
a wide range of behaviors, enabling researchers nationwide
to investigate the causes and progression of Alzheimer's
disease, brain aging, depression, and other conditions
— a brave new world of mouse-centric research.

Mice aren't perfect stand-ins for humans — for
instance, they don't have the same parts of the frontal
cortex that humans do, says Mikhail Pletnikov, an associate
professor of psychiatry and behavioral sciences at Hopkins
and one of the creators of the DISC-1 mouse, a murine
analog for human schizophrenia. ("Murine" is a term meaning
"of the mouse.") But, Pletnikov says, mice have become the
research subject of choice for scientists studying the
brain, especially during the last five years. "If you have
a gene, you can make a mouse," he says. "It's a tool that
scientists just have to use."

The march of
science has often been launched by big minds unlocking the
secrets of small things. Consider Democritus and the atom,
Leeuwenhoek and the protozoa, Pasteur and the bacterium,
Mendel and the gene.

In an allegedly ventilated, cage-lined cinder-block room in
the Ross Research Building, the small things are hardly
secretive. The smell — a potent mix of animal dander,
feces, and urine — is overpowering. "Most people
can't stand it, but I love it," says Keyata Thompson, a
research technologist for the School of Medicine's animal
research resources department. "It feels like home."

Thompson is one of about 90 people, including seven
veterinarians, who care for 200,000 or so mice at 10
facilities across Hopkins. The mice cost the university
about 75 percent of its annual $10 million animal care
budget — about 74 cents per day for each cage, an
average of about a quarter a mouse. (Cages typically can
accommodate five mice, but the Hopkins average is three.)
Why so many mice? Because they're cheap (about $2 to $3 for
a standard inbred mouse), manageable, portable, small, and
quickly bred. Using a single inbred line of mice —
ones created from a pair whose DNA has been specifically
altered by human hands — a viable cancer study can
take less than a year. With larger mammals, the timeline
grows exponentially.

Nearly a decade ago, 468 Hopkins faculty members kept
42,000 mice. Then things got crazy. "When I got here six
years ago, we were seriously overcrowded," says Julie
Watson, an assistant professor of molecular and comparative
pathobiology and the director of Hopkins' rodent medicine
and surgery program. "We've built new homes and taken
several new precautions to accommodate the mouse."

Because mice may carry germs and parasites, they can fall
prey to many illnesses — bad for the mice and
potentially for the caretakers. What's more, murine
ailments can also be bad for science. Reviewers of journal
articles will ask a researcher if his or her mice suffered
from health problems that could skew test results and,
hence, a paper's conclusions. "It's a question most
researchers really don't like to deal with," Thompson
says.

Several years ago, when Watson and leaders of the
university's animal research and resources department
realized the medical campus was being overrun by mice, they
looked with glee at the prospect of moving thousands of
them from rooms crammed with cages in older buildings into
a brand-new, state-of-the-art vivarium in the basement of
the Broadway Research Building (BRB), then under
construction. Too many mice were infected with
disease-causing pathogens, so Watson saw the new facility
as an opportunity to improve the quality of research.
"Infected colonies were infecting clean ones, so we needed
to come up with a plan," Watson says. "We told researchers
at all sites that they needed to clean up their mice if
they wanted to use this nice new facility."

Watson devised a program that took mouse pups away from
mothers who may be infected or infested. Investigators who
agreed to drop off their pups to Thompson and other animal
workers within 48 hours of birth for disinfecting
treatments wouldn't be charged for it. In return, they
could get their mice into BRB. Researchers bought into the
idea.

Now, besides making sure that mice at the medical campus
get plenty of rodent chow* — and
guaranteeing murine safety and "survivability" by demanding
that visitors and researchers wear head and shoe coverings
and smocks, and keep their perfume in the bottle at home
— Thompson also runs a production colony where she
breeds 10 to 20 mice per week. Because pathogens and worms
can be passed from generation to generation, Thompson takes
those mice from their mothers and cross-fosters them with
"clean," or germ-free, mothers. A so-called FBB mouse
specially bred to be incredibly maternal will wean mice of,
for example, the breed C57/black 6, a hardy staple of a
wide range of medical research.

A specially
made mouse
can
mimic
a purebred dog. "They're
expensive
and often
temperamental,"
Thompson says.

Before the pups are placed with their foster moms, they are
disinfected. Thompson uses chopsticks to remove the
grape-sized pups from their cages, places them in coffee
filters so they're easier to handle, then dunks the filter
into iodine for a few seconds. "It's nowhere near long
enough to drown them," she says. (Before Watson developed
this technique, standard procedures included Caesarian
sections and embryo transfers, which sacrificed the
mothers.)

When clean animals are moved to the BRB basement, they take
what Thompson calls "an animal transfer route" underneath
the medical campus. "People would freak out if you walked
through a facility with an animal," she explains.

You don't have to be anywhere near one of the 19 gleaming
"suites," each with five rooms, to sense that there are as
many as 100,000 mice in some 26,000 cages at BRB. All you
have to do is get on the elevator to experience the waft of
murine, um, essence. For humane reasons, mice are monitored
daily, with round-the-clock veterinary care for the sick.
At BRB and other facilities, mice enjoy fresh pumped-in
air, nesting materials that look like a towhead's matted
hair, corncob bedding, subdued lighting, and unlimited
feed. Robots douse and swab the cages every two weeks,
scrubbing away infections and worms. Stickers that read
OVERCROWDED are plastered on cages with more than five
mice, and researchers who don't fix the problem are charged
a daily fine. Of course, for all that comfort, a mouse's
lifespan — typically one to two years — might
still be shortened by research.

The university took some public relations hits in 2000,
when it was the lone research institution to lobby
successfully against more stringent federal regulations
targeted at university labs regarding mouse care. The
university argued that new rules, which would have required
research labs across the country to maintain detailed
paperwork on each individual mouse, would cost Hopkins
millions of dollars a year. So far, scientists who
sacrifice mice in the name of saving human lives have yet
to draw the same ire from animal rights activists as have
fur-wearers, meat-eaters, and vivisectionists. Most of the
complaints Hopkins receives regarding animal care have to
do with the School of Medicine's decision to continue to
use live pigs as surrogates during surgery. But there are
still worries. In February, extremists attacked the husband
of a breast cancer researcher at the University of
California at Santa Cruz because of his wife's use of lab
mice. As the use of larger mammals in research has waned,
often replaced by mice, activists have begun to take
notice, filling blogs with anecdotal instances of mouse
mistreatment and pointing to research that shows that as
much as 95 percent of all medical research is done using
mice or rats.

Hopkins researchers have developed more sensitive ways to
handle mice — and to avoid using them. Alan Goldberg,
director of the Johns
Hopkins Center for Alternatives to Animal Testing
(CAAT), notes how imaging devices that use nanotechnology
can show researchers how tumors develop and how therapies
attack them, without sacrificing the life of the mouse to
an autopsy. Much more research is being done in test tubes
instead of in living organisms, and there will likely be
more if federal strictures against human stem cell research
are loosened. Computer models in some cases are replacing
the mighty mouse. And scientists have increased the numbers
of non-mammalian species used in basic biological and
genetic research (good news for mice; not so good for fruit
flies and zebrafish).

Hopkins researchers have adopted many of those practices,
Goldberg says, but let's be clear: The outcome isn't always
pleasant for animals born into the service of humankind. At
some research institutions, more than 70 percent of male
mice are killed before weaning. That's not true at Hopkins,
but males that do not possess the right genes for
experimentation often are killed. That's the nature of the
research business.

The mouse's enduring utility as a research tool, even with
burgeoning alternatives, is a major reason for doing
everything to keep a mouse healthy. That, and price. While
basic inbred mice cost a couple of bucks, a knockout or
other genetically modified mouse can run in the tens of
thousands of dollars. In rare cases, an engineered mouse
can cost $100,000. An expensive mouse can temporarily
disappear into the wrong cage, causing a panic among
investigators until it is found and put back in its place.
There are other issues for handlers: A specially made mouse
can mimic a purebred dog. "They're the ones we usually
don't like to work with," Thompson says. "They're expensive
and often very temperamental." Apparently, a mouse of high
station can bite more.

Sometimes, the mice provide surprises, like the batch that
kept dying despite Thompson's care. The scientist hadn't
told her that the mice had been bred to have cardiac
problems. "I was mortified — I thought it was all my
fault," she says as she handles a mouse by the scruff of
its neck; the animal scrunches its eyes and makes noises
that can only be described as tiny.

Every six
months or so, a group of Hopkins pathologists, researchers,
and vets holds a conference that could be called "So, We
Got This Mouse. What Can We Do With It?" The Phenocore
Symposium is designed by vets to answer questions about
phenotyping — deciphering what value a mouse has to
science.

A phenotype is any observable characteristic that you can
measure, anything that distinguishes a mouse type. The
basic research mouse, like the black 6, may be better for
some experiments than others. Phenotyping can help
determine where it can be of most use. Even some transgenic
mice have identity issues. Researchers often aren't 100
percent sure how experiments should be designed around a
certain mouse, or why, in the worst-case scenario, a mouse
died — was it the experiment's design, or some
unforeseen genetic complication? At meetings and
conferences, Hopkins researchers from a wide variety of
specialties compare notes, offering tips on which mice can
be used for what.

Although genetics is key in phenotyping, environment can
also affect a mouse's viability as a research subject.
During a conference held at the medical campus in February,
Ellen J. Hess, associate professor of neurology and
neuroscience, recalled one group of mice that maintained
consistent activity levels in the lab all day long, then
quieted down at night, as is typical. But at around 2 a.m.
they went into a tizzy. Researchers were baffled by this
until a grad student slept over one night and discovered
that, precisely at 2 a.m., an investigator in the lab next
door cranked up his boom box, setting the mice to
dancing.

At the conference, speakers discussed blind mice, deaf
mice, mute mice, mice that "barber" the whiskers off of
other mice, and mice that died because they had been
implanted with human Alzheimer's genes and couldn't figure
out how to use a new water-delivery system in their cages.
The moral: Know thy mouse.

The conference also afforded a look inside the industry
that has sprung up to handle the mouse once it has been
bred, raised, and readied for research. On display were
six-lane treadmills, an assortment of mazes, a programmable
animal shocker, an eyeblink conditioning system, and the
Roto-Rod™" — a rotisserie-like device that
turns slowly in mid-air. Mice perch on the rod and turn
along with it, or fall off, indicating whether they are
viable for some types of research. At the dozen or so
exhibition tables set up outside Turner Auditorium could be
found the Vevo 770 Micro-Ultrasound System for housing and
testing mice — winner of a 2007 "Best Customer Value"
award. The 12-chamber CLAMS — for comprehensive lab
animal monitoring system — was touted by its maker,
Columbus Instruments, for keeping track of how much the
mice inside it have consumed and slept, their body mass,
ambient light and room temperature, their heart rate and
temperature, and other measurements. "They start at
$50,000," says Ken Kober, the company's sales manager, "and
then they start to get expensive. I've seen them go for as
much as $90,000. The Big Pharma guys love them."

Across the
city, at the Neurogenetics and Behavior Center at Mudd Hall
on the Homewood campus, even more specialized machines
— including odor discrimination equipment and
single-trial aversive learning cages — fill a
basement lab/vivarium. Here, researchers phenotype mice for
the benefit of researchers worldwide. The five-year-old
center has 40 Hopkins faculty members and several outside
partners, including the Jackson Laboratory and the Howard
Hughes Medical Institute. Backed by millions of dollars in
grants from the National Institutes of Health, the center
will put mice through specially designed paces to see what
they might bring to the brain-research table.

"We're trying to find the best ways to perform research on
the human brain using mice," says Michela Gallagher, vice
provost for academic affairs and a professor of psychology and
neuroscience at the Krieger School of Arts and
Sciences. "Do they function best in large groups? Will they
behave differently if they're removed from a group? We're
ground zero for getting those types of behavioral measures
out of mice."

The goal is to find ways to compare mouse and human
behavior, so that aging diseases like Alzheimer's and
mental illnesses like depression and schizophrenia can be
studied in rodents. For decades, psychologists have
preferred rats, often creating lesions in the animals'
brains to see how the lesions affected behavior. "Mice do
OK in our experiments, but they're not brilliant —
rats have been smarter," says Gallagher.

Yet, even here, in the one-time domain of King Rat, the
mouse reigns supreme. Because of its genetic malleability,
the mouse has become the animal of choice for the center's
faculty studying brain disorders. Instead of implanting
lesions, or even genes, in rats, researchers have made
transgenic mice their instrument of choice because of their
availability, and Hopkins researchers' ability to create
and rapidly breed them.

Research technologists disinfect grape-sized mouse pups,
then place them with foster mothers.

The pre-eminence of mice has changed brain science. "The
mouse represents a new phase in my research," says
Pletnikov, a member of the Neurogenetics and Behavior
Center. "I've had to learn how to do genetic models —
to insert DNA into a mouse genome. Previously, I
concentrated entirely on how viruses and other
environmental factors affect the brain. Now I'm also
investigating how proteins and receptors — the
molecular pathways that can affect brain health — are
regulated by genes." Mice implanted with the DISC-1 gene
show the same hyperactivity and abnormalities in social
behaviors as humans who suffer from the disease — a
sign that they have validity as research tools for
investigating schizophrenia in humans, Pletnikov says.

Disorders such as schizophrenia are present in a person's
genes before birth, but only manifest themselves after the
onset of adulthood. Mice can add an extra dimension for
researchers because of recent discoveries that allow for
controlling a gene's expression with the help of
pharmacology. For example, researchers can "turn off" a
gene or delay its expression by giving a young mouse a
simple drug in its food (in this case, the common
antibiotic tetracycline). The drug binds to a protein that
prevents the protein from binding to another one upstream,
an event along a so-called molecular pathway that would
activate the schizophrenia gene. Once the drug is removed,
the adult mouse will demonstrate the murine equivalent of
human schizophrenic behavior. Because the mouse can be both
a disease carrier and yet not exhibit behavior of the
disease, it accurately approximates the manifestation of
schizophrenia in people. The mouse also performs double
duty for researchers because they can see how the gene acts
when it is on or off. "The control is built right into the
mouse," Pletnikov says.

In the basement at Mudd, Gallagher and her crew study
aging, autism, eating and obesity, learning, and memory in
mice while searching for ways to compare mouse behavior
with that of humans who suffer from disease. For example,
Gallagher is working to model the onset of Alzheimer's.
Brain circuits in rodents with the disease that govern
their spatial memory appear to break down early —
something that can be extrapolated to future studies of
human subjects, she says.

The potential for so-called translational work is what
drives the center, says Alex Johnson, a post-doctoral
fellow in psychological
and brain sciences. "We're doing things with mouse
genetics that no one else is doing," he says.

Science has
not yet exhausted the uses of the humble mouse — far
from it. For example, as good as mice have been for his
cancer studies, Pardoll says the future might be brighter.
The advent of the "humanized mouse" — one that
carries a match of the tumor and the blood system,
including the immune system, of a particular patient
— is on the horizon. That may lead to the ultimate
mouse model: one unique to an individual person's biology.
Research so far is promising, but is hung up on two
questions: Will the tissue of a mouse reject the human
immune system? And, human blood travels through arteries,
capillaries, and vessels, but unlike in a mouse's system,
it also flows through certain cell membranes. Will a
mouse's biology be able to handle it? "It all seems
possible, but doesn't quite add up yet," says Pardoll.
"We're many years away from answering those questions."

Such research will continue as the prospects of the mouse
skitter off in some bold, often opposite directions. The
National Institutes of Health is pouring $60 million into a
"knockout mouse project" to map the function of every known
mouse gene to aid the cause of ever-more-specific research.
At the same time, the federal government and the Jackson
Laboratory are breeding 1,000 new and genetically mixed
mouse strains from the original eight basic inbred ones. A
more varied population of mice than the descendants of C.
C. Little's should create genetic diversity — all the
better for them to represent the diversity of humans in
experiments. Some scientists blame faulty — and
ultimately deadly — clinical trials involving such
drugs as Vioxx and Phen-fen on the fact that they were
initially performed on mice with the same handful of genes,
yielding poor translational results. Others argue that
medical research hasn't gone as far as it could because the
mice and their gene pools haven't been up to it.

Another school of thought believes that mice confined in
dark silence do not develop fully, further limiting their
suitability as research subjects. A mouse's surroundings,
the thinking goes, should be made more stimulating, so
their brains and nervous systems can flourish, or more
closely approximate what they would become in nature.
"Geneticists have thought of the mouse as a furry test
tube," says Cory M. Brayton, an associate professor of
comparative and molecular pathobiology and director of the
university's phenotyping program. "You drop your mutated
gene in there and you should find out specifically what it
does. They forget that the mouse is an animal with a
biological system."

Such thoughts don't veer all that close to
anthropomorphism, but they do show that an animal drafted
as a stand-in for humans might just be worthy of a morsel
of respect.

Michael Anft is a senior writer for Johns Hopkins
Magazine.

___________________* This corrects earlier information published in the
September issue print edition.